Approximately 7.2% of adults in the United States in 2012 suffered from an Alcohol Use Disorder, as defined by the American Psychiatric Association. Alcohol is responsible for a number of severe diseases, including alcoholic liver disease, alcohol related dementia and fetal alcohol syndrome. Past research has focused on organs such as the liver, or brain, but it has become increasingly evident that alcohol has significant effects on nearly all tissue in the body. One tissue that has garnered recent attention in part due to the rapid rise in obesity, is adipose tissue. Recent studies have provided compelling evidence relating the dysfunction of adipose tissue to the progression of many diseases, including liver disease. The interaction between ethanol and adipose tissue, particularly brown adipose tissue, has been understudied. There is little known about how ethanol impacts brown adipose tissue function, and further understanding of this relationship may prove crucial in elucidating origins of systemic changes seen in chronic alcoholics. This proposal will investigate the effects of ethanol on brown adipocyte development and function, elucidate mechanisms involved, and study the consequences of these effects on metabolic homeostasis. Both an in vitro and an in vivo model will be used. An in vitro model will be used to investigate whether ethanol interferes with the development of brown adipocytes. Brown preadipocytes isolated from wild-type C57BL/6J mice will be differentiated into mature brown adipocytes with or without exposure to 100mM ethanol. RNA and protein isolates will be collected from the differentiated cells. Markers of adipocyte character and function will be evaluated by qRT-PCR and western blot. Additionally, proteins involved in the mTOR pathway will be evaluated via western blot and immunoprecipitation to elucidate mechanisms that may be involved with brown adipocyte function. A similar approach will be applied to mature brown adipocytes. These studies are the focus of specific aims 1 and 2. An in vivo model will be used to study the effects of chronic ethanol intake on brown adipose tissue function, and to relate these effects with whole body metabolism. Wild-type C57BL/6J mice will be given a chronic ethanol diet. Groups of mice will be exposed to conditions that have been shown to mediate brown adipose tissue activity, including cold habitat, beta adrenergic injection, high fat diet (al 3 increase BAT activity), and thermoneutral habitat (decrease BAT activity). Weight and temperature will be recorded throughout the study, and mice will be sacrificed for collection of tissue. Various depots of adipose tissue will be collected, along with liver and muscle. These tissues will be evaluated for both functional markers, and markers of brown/beige/white adipocyte fate. These experiments are the focus of specific aim 3.